Midterm 3

Question 1

1. Understand the relationship between ascorbic acid, dehydroascorbate, and 2, 3-diketogulonic acid

Answer 1

Ascorbic acid→ ascorbate free radical (Lost one electron) → dehydroascorbate (Lost 2 electron) → 2,3-diketogulonic acid (inactive) (Irreversibly oxidized)..

Question 2

2. What are the differences in the absorption of ascorbic acid and dehydroascorbate, which form is better absorbed? How is it transported in blood/plasma?

Answer 2

a. Ascorbate is absorbed by a Na-dependent, saturable, active transport system; ascorbate may be oxidized to dehydroascorbate and absorbed by GLUT1 or GLUT3.
b. Dehydroascorbate appears to be better absorbed; once absorbed, it is rapidly reduced back to ascorbate by ascorbate reductase involving NADPH.
c. Up to 200 mg is absorbed, but absorption at higher doses (>10g) is limited (<20%), and may result in malabsorption and lead to osmotic diarrhea.
d. Once in the enterocyte, vitamin C is transported across the basolateral membrane by Na-independent facilitated transport, and exist in plasma mainly as free ascorbate.

Question 3

3. What are the major co-enzyme functions associated with vitamin C; what are the mechanisms in these functions; what is the non-coenzyme function, why is the general antioxidant activity of vitamin C not considered a vitamin function?

Answer 3

a. Vitamin functions – The vitamin roles of ascorbate are defined as its functions that are required in the prevention of scurvy; it acts as a reducing agent, and in many cases, it is involved in hydroxylases in hydroxylation reactions of various substrates.
i. Collagen synthesis – Ascorbate is important in the post-translational hydroxylation of proline to hydroxyproline, and lysine to hydroxylysine. Hydroxyproline is important in the formation of the triple-helical structure of collagen; hydroxylysine is important in the cross-linking of collagen fibers. Ascorbate, in these reactions, is specifically involved in the reduction of ferric iron to ferrous iron.
1. Proline → Hydroxyproline
2. Lysine → Hydroxylysine
ii. Carnitine synthesis – Ascorbate is important in two reactions in the conversion of trimethyllysine to carnitine. The first step involves the hydroxylation of trimethyllysine to 3-OH trimethyllysine by trimethyllysine hydroxylase; the final reaction is the conversion of 4-butyrobetaine to carnitine by 4-butyrobetaine hydroxylase – both these enzymes are vitamin C dependent.
1. Trimethyllysine → trimethyllysine hydroxylase → 3-OH-trimethyllysine
2. 4-butyrobetaine → 4-butyrobetaine hydroxylase → Carnitine
iii. Tyrosine synthesis & metabolism –
1. Ascorbate is involved in the conversion of phenylalanine to tyrosine by phenylalanine hydroxylase.
2. Other vitamin C dependent, tyrosine-related enzymatic reactions include: P-hydroxyphenylpyrubate → p-hydroxyphenylpyruvate hydroxylase → homogenitase
3. Homogentisate → homogentisate → 4-maleylacetoacetate.
iv. Cholesterol metabolism – The hydroxylation of cholesterol to 7-OH cholesterol by cholesterol a-7 hydroxylase; impaired function of this enzyme results in decreased bile synthesis and elevated cholesterol.
b. Non-vitamin functions – Plays a role as a general antioxidant in aqueous phases, but is not considered a vitamin function, as it can be substituted by other antioxidants; increases in non-heme iron absorption (can be done by other acids).

Question 4

4. What is the estimated total body pool size for ascorbate; how much dietary ascorbate is needed to maintain this pool, how are mega doses of ascorbate handled by the body?

Answer 4

a. The body pool is believed to be between 1,500 – 3,000 mg
b. This can be maintained by 100 – 200 mg/day
c. Concentrations are highest in the organs.
d. Increasing ascorbate intake (mega doses) does not appear to increase urinary oxalate excretion and large amounts of dietary/supplemented ascorbate appears to be excreted unmetabolized as free ascorbate in urine.

Question 5

5. What is the mechanism of ascorbate’s enzymatic role, how are Fe and Cu involved?

Answer 5

a. Ascorbate’s enzymatic role: both iron and copper are important in several hydroxylation reactions responsible for ascorbate’s major functions. Ascorbate is also important in the absorption of non-heme iron and facilitates its absorption by maintaining it in the reduced form (Fe+2).
b. Alternatively it may increase elemental iron absorption by forming a soluble complex with ascorbate. Ascorbate is also important in the distribution and mobilization of iron stores within the body.
c. Absorbate is important in getting iron in the proper form to be released from ferritin and transferrin molecules.
d. Absorbate is also thought to interfere with the movement of copper into its transport protein ceruloplasmin. Ascorbate supplementation of 1.5 to 2 g/ day resulted in lower plasma concentrations of copper as ceruloplasmin

Question 6

6. What is the importance of ascorbate in the amidation of peptides with C-terminal glycine residues (be able to cite a few of the important products)?

Answer 6

a. Ascorbate is important in keeping Cu in its proper form (Cu+) in the Cu-dependent enzyme peptidylglycine alpha-amidating monooxygenase. In this reaction the C terminal glycine of polypeptides is cleaved between its amino group and the central alpha carbon leaving the C-terminal end of the peptide amidated.
b. The products are important neurotransmitters, hormones, and hormone releasing factors such as CCK, gastrin, oxytocin, calcitonin, bombesin, corticotropin releasing factor, and growth hormone releasing factor.

Question 7

7. What is the deficiency disease associated with vitamin C, what are some of the symptoms, how might these be related to its functions?

Answer 7

a. Scurvy – This results when the total body pool of ascorbate falls below 300 mg; it is estimated that as little as 10 mg/day would prevent the symptoms of scurvy.
b. 4H’s represent the symptoms of scurvy – hemorrhage, hyperkeratosis, hypochondriasis, and hematologic.

Question 8

8. What are the potential toxicities associated with mega doses of ascorbate; to what extent do these represent toxicities? (be able to support your opinion)

Answer 8

a. Ascorbate is poorly absorbed at large doses (>1g), and may cause osmotic diarrhea.
b. A putative toxicity reported is the occurrence of kidney stones (oxalate stones) in individuals who are susceptible. Nevertheless, there is no biochemical/metabolic evidence

Question 9

9. What environmental factor is important in assessing an individual’s requirement for vitamin C?

Answer 9

a. Individuals who smoke have a greater need for ascorbate due to the destruction of the vitamin by oxidative products of smoking. The estimated increase for smokers is an additional 35 mg/day.

Question 10

10. What are the major association between vitamin C and chronic disease; what problems are associated with acceptance of these interpretations?

Answer 10

a. Common cold – There is no evidence that vitamin C prevents the common cold; nevertheless, there is some evidence to indicate that vitamin C reduces the symptoms/durations of the cold by acting as an anti-histamine at higher doses, with equal evidence that indicates otherwise.
b. Cancer – vitamin C can potentially prevent cancer development, as evidenced by in vitro and animal studies; evidence is inconclusive in humans, and potential evidence that vitamin C protects against cancer is confounded with high fruit/vegetable intake. Vitamin C may have an important role in gastric and colon cancers.

Question 11

1. What is the oxidation state of Ca; what are the major inorganic forms?

Answer 11

a. Oxidation state: Ca+2

b. Major inorganic forms: ionized: Ca+2, calcium phosphate: Ca3(PO4)2, hydroxyapatite: Ca10(PO4)6(OH)2

Question 12

2. What is the estimated body content, how is this distributed in the body; what are intracellular and extracellular concentrations, what regulates this distribution of Ca in the body?

Answer 12

a. Estimated body content: 14,000,000 ppb; 980-1000g
b. Distribution: 99% of the calcium is in the skeletal system with the remaining 1% distributed in intra and extracellular compartment
c. Intracellular and extracellular concentrations: intracellular concentrations of ionized Ca are maintained at very low levels about 0.1uM, while extracellular concentrations of ionized calcium are approx. 2-2.6 mM
d. Regulation of distribution:
i. Calbindin system- the system is regulated by need and can increase Ca absorption from an average of 30% to as much as 60-70%.
ii. Non-cellular transport- operates primarily in the distal jejunum and ileum and is paracellular in nature. This mechanism is non-saturable and non-carrier mediated and dependent solely on concentration gradient.
iii. Parathyroid hormone and gland regulate the concentration of calcium in the plasma
iv. If increased absorption is not enough, vitamin D will stimulate the differentiation of monocytic stem cells into mature osteoclasts. The osteoclast will cause the removal of calcium and phosphorous from the bone.

Question 13

3. What are major dietary sources, and what influences the bioavailability of Ca?

Answer 13

a. Dietary sources: dairy products including milk, cheese, yogurt, and ice cream, seafood
b. Influences of bioavailability: an acid pH favor Ca absorption, while at alkaline pH as is found in small intestine, Ca may complex with other small ions and food constituents that reduce its availability

Question 14

4. How is Ca absorption regulated hormonally, what is the average absorption from foods and supplements?

Answer 14

a. Calcium from foods and supplements are absorbed at a 30% rate under normal conditions.
b. There are two major transport systems responsible for Ca absorption.
i. Calbindin system: Active transport system that is regulated by 1,25 (OH)2-D; active when Ca intake is low or as a result of a fall in serum Ca. Absorption is regulated based on need and solubility – Calbindin can increase Ca absorption to 60-70% Vitamin D co-ingestion with calcium can significantly increase absorption.
ii. Ca/Mg ATPase pump in the proximal small intestines; Na/Ca ATPase pump in the distal small intestines.
iii. Non-cellular transport – non-saturable and non-carrier mediated and dependent on concentration gradient.
iv. Colonic absorption

Question 15

5. How is Ca involved in bone mineralization and remodeling?

Answer 15

a. 99% of Ca is present in bone is found as crystalline salts – calcium phosphate, hydroxyapatite.
b. Osteoblasts secrete several proteins which bind calcium to the surface of the bone at the site mineralization until the critical concentration of Ca, PO4, and mg are met, thus beginning the crystallization of the salts.

Question 16

6. What are the major non-osseous roles for Ca?

Answer 16

a. Of the 1% of Ca not found in bone;
b. Enzyme activation/deactivation
c. Changes in membrane permeability
d. Activation of clotting factors
e. One function is in enzyme activation/deactivation – covalent modification.
i. Protein kinase C will activate this enzyme, causing phosphorylation.
ii. Ca/Mg ATPase (+)
iii. Glycerol-3- phosphate dehydrogenase.
iv. Pyruvate kinase (– in kidney and liver)
v. Pyruvate carboxylase (+ influence in liver and kidney)
vi. Pyruvate dehydrogenase (–)
vii. Glycogen synthase (–)
viii. Glucose-6- phosphatase (+)
f. Muscle contractions.
g. Microfibril contractions.
h. Cellular secretion (i.e. insulin secretion from Beta-cells).
i. Activation of clotting factors.
j. Nerve conduction.

Question 17

7. What are the important interactions between Ca and other nutrients in digestion and absorption, and in bone mineralization?

Answer 17

a. Ca absorption is interfered with by excess Mg, Fe, Zn, P, fatty acids, proteins, and phytates.
b. Ca decreases heme and non-heme iron absorption.
c. Sodium increases Ca excretion; potassium decreases calcium excretion.
d. Dietary protein increases Ca absorption as well as increasing Ca excretion; high protein diets are thought to increase risk of premature osteoporosis – evidence for this (?)
e. Rena calcium reabsorption is controlled hormonally by PTH>
f. Calcium is excreted through urine and feces, not sweat.

Question 18

8. What is the relationship between Ca and development of osteoporosis; what factors influences accrual of bone minerals over an individual’s lifespan; what is meant by the statement: osteoporosis is a disease of the youth, manifested in old age?

Answer 18

a. Bone is in a continual state of turnover, and being remodeled. The renewal rate is ~100% in infant and 18-20%/yr in adults.
b. Bone mineral accumulates during the early part of our lives and reaches a peak between 25-35 age,10-15 yrs after cessation of linear growth. Afterwards, bone resorption exceeds bone deposition and bone loss occur, resulting in the loss of bone mineral and bone protein content.
c. This slow and insidious process of bone loss is accompanied with changes in the micro-architectural structure, thinning of both cortical and trabecular bone – developing osteoporosis.
d. Higher peak bone density results in the delaying of osteoporotic level – important to accumulate bone
minerals during the critical years.

Question 19

9. What is the importance of gravity on bone mineralization?

Answer 19

a. Osteoporosis is defined as “a systematic skeletal disease characterized by low bone mass and microarchitectural deterioration with consequent increase in bone fragility and susceptibility to fracture” by the NIH.
i. Type I – Rapid bone loss, primary trabecular, mostly in women post-menopausal 51-65 yrs. Increased compression of bone and loss of height.
ii. Type II – Slow loss of both cortical and trabecular bone, both men and women > 75 yrs.
b. Importance is placed on the younger years (10-15) – this results in greater peak bone density. Individuals with highest peak bone mass at adolescence have the greatest protective advantage against the development of osteoporosis.
c. However, milk consumption in youth has declined over the past few years.
d. Gravity is constantly putting pressure on our bones which causes our body to add bone mass into our bone which causes a stronger bone.

Question 20

10. You should be able to explain the process of bone mineralization and remodeling in general terms (this includes integrating what you have learned about the importance of other nutrients in the process)

Answer 20

a. Parathyroid hormone is released by the parathyroid gland which stimulate the conversion of 25-OH vitamin D to 1, 25 (OH)2 Vitamin D. 1, 25 (OH)2 Vitamin D stimulates the differentiation of monocytes into osteoclast which is responsible for the removal of calicum in the bone. They also form osteoblasts which the mineralization of bone.

Question 21

1. What is the oxidation state of P; what are the major inorganic forms?

Answer 21

a. Oxidation state: -3, +3, +5

b. Major inorganic forms: amorphous calcium phosphate, magnesium phosphate, hydroxyapatite.

Question 22

2. What is the estimated body content, how is this distributed in the body; what are its important intracellular roles?

Answer 22

a. A 70 kg individual contains between 700-770 g of phosphorous – 85% is found in bone as calcium phosphate and hydroxyapatite; the rest (15%) is distributed in soft tissue – most present in the form of phospholipids of cell membranes.
b. Most phosphorous is free filterable phosphate, while 10% is bound to protein, and 5% bound to Ca and Mg.
c. Inorganic phosphate is important in that it acts a buffer.

Question 23

3. What are major dietary sources; what are the major forms; how is it absorbed; how is this regulated? (you should be able to answer this in very general terms)

Answer 23

a. Phosphorous is widely distributed, particularly in animal products, including meats, fish, poultry, eggs,and milk, as organophosphates. Plant sources include nuts and whole grains, although its bioavailability from whole grains is low because much of it is bound phytate, and thus unavailable for absorption.

Question 24

4. How is P involved in bone mineralization and remodeling?

Answer 24

a. 85% of phosphorous is found in bone as amorphous calcium phosphate, magnesium phosphate or hydroxyapetite.

Question 25

5. What are the major non-osseous roles for P?

Answer 25

a. Component of cell membranes (phospholipids)
b. Intracellular buffers
c. High-energy phosphate bonds (ATP)
d. Cell signaling
e. Metabolic trapping (phosphorylation)
f. Enzyme regulation (covalent modification involves phosphorylation/dephosphorylation).
g. Activation of B vitamins (PL + P i PLP)
h. Component of genetic material (DNA & RNA).

Question 26

6. How is P excreted from the body, in what way is this regulated?

Answer 26

a. Increase in dietary P results in an increase in phosphorous excretion.
b. Approximately 1/3rds of phosphorous is excreted in feces; the rest in urine.

Question 27

1. What is the oxidation state of Mg; what are the major inorganic forms?

Answer 27

a. Oxidation state: +2

b. Major inorganic forms: Mg+2

Question 28

2. What is the estimated body content, how is this distributed in the body; what regulates this distribution of Mg in the body?

Answer 28

a. 65-70% in bone
b. 25-30% in muscle
c. 6-7% in other cells
d. 1% extracellularly
e. Regulation by absorption, excretion and transmembrane flux rather than hormonally.

Question 29

3. What are major dietary sources, and what influences the bioavailability of Mg?

Answer 29

a. It is found in a wide variety of foods including whole grains, nuts, legumes, teas, coffee, and cocoa.

Question 30

5. What are the major non-osseous roles for Mg? (this is very general answer)

Answer 30

a. The majority of Mg is found in the free ionized form intracellularly (35-40%) and is generally associated with phospholipids on membrane surfaces complexed with DNA, proteins, and ADP/ATP.
b. Mg is a component of metaloenzymes, where it is a component of ATP, and to a lesser extent, GTP.
Stabilize the phosphate ester bonds and facilitates various intracellular phosphorylation / dephosphorylation reaction

Question 31

1. What is the estimated total body water, how is it calculated; how is this estimate influenced by body composition; what is the estimated volume distribution (%) of intracellular and extracellular water?

Answer 31

a. Total body water (TBW) can be calculated by isotopic dilution using stable isotopes. It is estimated in normal healthy adult males from the following formula assuming TBW represent 54-55% of body weight.
b. Women and obese individuals – greater percent body fat and would have a lower percent TBW than predicted by body weight alone.
c. Body surface area – infants have greater surface area and therefore, fractional water volume is greater in infants.
d. Estimated volume distribution:
i. Intracellular fluid volume- 60%
ii. Extracellular fluid volume- 40%
1. Plasma 8%, interstitial 28%, transcellular 4%

Question 32

2. What is the difference between plasma water and plasma fluid concentrations?

Answer 32

a. Plasma fluid – all fluids in plasma; plasma water – the water in plasma.

Question 33

3. What is the difference between osmolality and osmolarity; what are effective and ineffective osmols?

Answer 33

a. 1 osmolarity is volume dependent
b. 1 osmolality is weight dependent
c. Solutes that can equilibrate across membranes such as urea and alcohol will simply move down a concentration gradient to establish and maintain osmolality. These solutes are called ineffective osmols since they do not generally effect water movement.
d. Changes in solute concentrations that do not move freely across membranes (effective osmoles) such as Na+ and glucose will cause water to move from intracellular space to extracellular space in order to equilibrate solute concentrations.

Question 34

4. You should know the relative intracellular/extracellular distribution of Ca, P, Mg, Na, K, Cl and bicarbonate (in other words, is the solute primarily intracellular or extracellular?).

Answer 34

a. Ca - Extracellular: 2.0-2.5mM, Intracellular: 10^-4 mM
b. P -
c. Mg - Extracellular: 1.8-3.0mM, Intracellular: 3.5mM
d. Na – Extracellular: 136-144 mM, Intracellular: 12 mM, about 100g total in the body – 40% of which is in bone, and 60% in extracellular fluid;
e. Cl – Extracellular: 110 mM, Intracellular: 2 mM;
f. Potassium – Extracellular: 4-5 mM, Intracellular: 150 mM, its presence intracellular fluids is 30-40 fold higher than in extracellular fluids.

Question 35

5. What are the oxidation states of Na, K, and Cl?

Answer 35

a. Na +
b. K +
c. Cl -

Question 36

6. What are the general dietary sources of Na, K, and Cl; what percentages of these dietary electrolytes are absorbed? How might potassium present in unprocessed foods differ from potassium present in processed foods or salt substitute (KCl)?

Answer 36

a. Na & Cl – Animal products are good sources of sodium; 90% of sodium intake comes in the form of NaCl, with 15% from added table salt, and 75% from salt added during the processing foods.
b. K – Fruits and vegetables are good sources of potassium. Potassium from natural sources are balanced by citrate or other conjugate anionic bases, which can be converted in cells to a bicarbonate anion which acts as a buffer. In contrast, potassium from processed foods is added with a chloride salt and provide little/no buffering capacity.
c. Over 90% of dietary sodium, potassium, and chloride and nearly all of the sodium and chloride secreted into the lumen of the small intestine is absorbed.

Question 37

7. What are the major functions of Na, K, and Cl?

Answer 37

a. Na –
i. Maintenance of fluid volume and plasma osmolality
ii. Maintenance of membrane potential
iii. Action potential in muscle and nerve cells
iv. Membrane nutrient transport systems
v. Enzyme catalyzed reactions: Na/K ATPase pump
vi. Sodium is responsible for maintaining osmotic and ionic balance in extracellular fluid, as well as the electrochemical gradient in nerve cells, and participating in acid/base balance. Sodium concentration is tightly regulated by aldosterone, an adrenal steroid hormone.
b. K –
i.
ii. Membrane nutrient transport systems
iii. Enzyme catalyzed reaction: Na/K ATPase pump and pyruvate kinase
iv. Acid/Base Balance: balancing the charges of intracellular proteins, inorganic, and organic phosphates
c. Cl –
i. Enzyme catalyzed reaction: Cl is needed for the activity of angiotensin-converting enzymes
ii. Chloride shift: maintaining extracellular bicarbonate concentration
iii. Important for the synthesis for HCL acid in gastric mucosa

Question 38

8. What is the relationship between Na and K and the development of hypertension; between Ca and Na and hypertension?

Answer 38

a. Increasing potassium intake, rather than decreasing sodium intake, seems to exert greater effects in reducing hypertension in most individuals. This is due to the fact that sodium concentrations are tightly regulated by homeostatic mechanisms; hence, large variations in sodium intake do not seem to influence plasma sodium concentrations to any great extent.

Question 39

9. What is the major cause of hyponatremia?

Answer 39

a. Accumulation of water and impaired water excretion; the inappropriate loss of sodium relative to water, or excessive water consumption.